Zinc deficiency contributes significantly to world wide burden of disease accounting for 10-15% of morbidity and mortality associated with diarrheal disorders and pneumonia. Physiological hypozincemia of aging may account for sensitivity of this population to lower respiratory tract infections. From a pragmatic point of view: a) Zn is involved in innate, adaptive and nutritional immunity; and b) Zn supplementation is inexpensive and has a relatively large therapeutic index. Nonetheless, there is a notable lack of preclinical assessment of mechanisms and therapeutic efficacy of supplemental zinc in the context of pandemic flu (H1N1) with or without secondary bacterial infection (Methicillin-resistant S. aureus, MRSA). In preliminary experiments, we noted that dietary zinc deficiency (Zn-D) did not affect the sensitivity of intact mice to either H1N1 or MRSA, but exposure to both (H1N1/MSRA) resulted in more severe acute lung injury (ALI). Further preliminary data showing that cell death (apoptosis) of distal airway epithelium and inability of alveolar macrophages to phagocytize apoptotic cells (efferocytosis) in Zn-D suggest that mechanistic insight may be obtained in co- culture model system of human alveolar type II cells (ATII) and alveolar macrophages (AM). Additional preliminary data showing that aged mice are Zn-D and sensitive to H1N1 suggest that this is a useful model to consider pharmacotherapeutic potential of Zn. Accordingly, SPECIFIC AIMS are: 1.Determine role of Zn dyshomeostasis in ALI due to H1N1/MRSA. The sensitivity of distal lung to combined infection will be assessed in mice made Zn-D and contrasted to zinc replete (Zn-R) controls. The ability of supplemental Zn to modify injury to and recovery from H1N1/MRSA will be assessed by correcting Zn deficiency prior (preventive) to or after (mitigative) exposure. 2. Determine the molecular mechanisms by which Zn modifies inflammatory response to viral and bacterial infection in cells of distal lung. We will determine: ) if Zn deficiency affects human ATII cells and AM response to H1N1/MRSA; b) the mechanism by which GM-CSF and TGFß affect Zn homeostasis and cellular function in human AM; and c) if Zn supplementation and NO affect human ATII and AM response to H1N1, MRSA or H1N1/MRSA. 3. Determine therapeutic efficacy of Zn in physiological hypozincemia of aging and sensitivity to viral and bacterial infection. Aged (18-22 months) mice will be made Zn replete and the effect of H1N1/MRSA will be assessed. A separate cohort of aged mice will be infected with H1N1/MRSA and intrapulmonary labile levels of zinc will be elevated by inhaled nitrite. Collectively, these studies will provide novel insights into the role of Zn in regulating resolutio of H1N1/MRSA-induced lung injury through modulation of alveolar epithelial apoptosis and macrophage efferocytosis. Such mechanistic insight may help guide the use of zinc in: a) preventive fashion for global health issues regarding sensitive populations to viral and bacterial pneumonia; and b) suggest combined therapies of zinc and nitrite to mitigate pneumonia in aged population.

Public Health Relevance Statement: Acute lung injury following viral and bacterial infections continues to be a global public health problem and results in significant mortality and morbidity. Despite advances in the understanding of the mechanisms that govern acute infections, discovery of new and efficacious therapeutic agents for the resolution of acute lung injury has lagged behind. In this proposal, we will explore the role of zinc as a crucial regulator of resolution phase of acute lung injury following H1N1 and MRSA lung infections.